Paraffin-embedded tissue as a source of RNA for gene expression analysis in oral malignancy.
OBJECTIVE
To assess the feasibility of using archival oral mucosal tissue to examine gene expression at the ribonucleic acid (RNA) level.


METHODS
We describe the isolation of RNA from 8 microns sections of formalin-fixed paraffin-embedded oral mucosal tissue. RNA was reverse transcribed and three candidate genes amplified by polymerase chain reaction (PCR). The ribosomal protein S14 gene is a housekeeping gene which has been used as an internal standard in several quantitative PCR protocols. The thymidine kinase (TK) gene is expressed at low levels in most tissues and, with a well-documented genomic organisation, is a useful tool for discrimination between genomic DNA and cDNA. The RI alpha gene is reported to be overexpressed in many cancer cell lines, in malignant tissue and in vitro transformed cells.


RESULTS
The S14 gene, the TK gene and the RI alpha gene of the cAMP-dependent protein kinase (PKA) were amplified successfully from formalin-fixed paraffin-embedded tissue sections. The TK primer pair is a useful additional tool in the unambiguous identification of RNA-derived species.


CONCLUSIONS
RNA suitable for reverse transcribed (RT)-PCR was extracted from archival oral mucosal tissue. This should permit rapid sequence analysis of transcribed tumor suppressor genes and oncogenes in this material. Furthermore, the RT-PCR approach described may allow quantification of gene expression in oral mucosal archival material processed in a standard fashion.
Cairns
M T
MT
Department of Oncology, Queen's University of Belfast, Belfast City Hospital Tower, UK.
Church
S
S
Johnston
P G
PG
Phenix
K V
KV
Marley
J J
JJ
D002294:Carcinoma, Squamous Cell / Q000235:genetics*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics; D005260:Female; D015870:Gene Expression; D006801:Humans; D008875:Middle Aged; D009857:Oncogenes / Q000235:genetics*; D016612:Paraffin Embedding; D010865:Pilot Projects; D016133:Polymerase Chain Reaction / Q000379:methods; D011505:Protein-Tyrosine Kinases / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis / Q000302:isolation & purification*; D012334:RNA, Neoplasm / Q000032:analysis / Q000302:isolation & purification*; D012194:RNA-Directed DNA Polymerase; D012269:Ribosomal Proteins / Q000235:genetics; D014062:Tongue Neoplasms / Q000235:genetics*
D012333:RNA, Messenger; D012334:RNA, Neoplasm; D012269:Ribosomal Proteins; C039747:ribosomal protein S14; D011505:Protein-Tyrosine Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D012194:RNA-Directed DNA Polymerase